Linker histone H1-0 is a specific mediator of the ETV6::RUNX1 transcriptional landscape in preleukemia and B cell acute lymphoblastic leukemia [RNA-seq]

The chimeric hematopoietic transcription factor ETV6::RUNX1 is the most common oncogenic fusion in pediatric B cell precursor acute lymphoblastic leukemia (BCP-ALL). It induces a clinically silent preleukemic state that requires secondary mutations for progression to full-blown leukemia. ETV6::RUNX1 functions primarily through repression of RUNX1 binding sites. In order to elucidate the characteristic quiescent state of ETV6::RUNX1 expressing cells, we generated preleukemic human induced pluripotent stem cell (hiPSC) models from two healthy donors using CRISPR/Cas9 gene editing. We identified upregulation of linker histone H1-0 in preleukemic hiPSCs, which was preserved upon hematopoietic differentiation and transformation to overt BCP-ALL. ETV6::RUNX1 induces activity of the H1-0 promoter whereas depletion of H1-0 specifically inhibited ETV6::RUNX1 signature genes, indicating its role as a novel key mediator of the quiescent ETV6::RUNX1 transcriptional profile. Single-cell gene expression analysis revealed that H1-0 levels strongly anti-correlate with cellular transcriptional activity, resulting in particularly high expression in quiescent cells during hematopoietic development. Pharmacologically, H1-0 protein levels correspond to susceptibility of BCP-ALL towards histone deacetylase inhibitor (HDACi) treatment and our data indicate efficacy of combinatorial drug treatment using the potent H1-0-inducing HDACi Quisinostat in BCP-ALL expressing high basal H1-0 levels. Overall design: To analyze the transcriptional landscape conferred by the ETV6::RUNX1 fusion gene, we generated knock-in human induced pluripotent stem cells (hiPSCs) derived from two healthy donors (HW8 and ChiPSC12). In the generated hiPSC lines (n=3), ETV6::RUNX1 is expressed under control of the endogenous ETV6 promoter. We performed gene expression profiling analysis using data obtained by RNA-seq of 2 wild-type hiPSC lines (HW8 WT and ChiPSC12 WT) and 3 monoclonal ETV6::RUNX1+ hiPSC lines (HW8 E::R 1, HW8 E::R 2, ChiPSC12 E::R). To determine the contribution of H1-0 to ETV6::RUNX1+ BCP-ALL pathology, we knocked down H1-0 in the ETV6::RUNX1+ BCP-ALL cell line REH using siRNA pools. We performed gene expression profiling analysis using data obtained by RNA-seq of REH cells treated with a non-targeting siRNA pool (siCtrl) and two H1-0-targeting siRNA pools (siH1-0_1, siH1-0_2) for 48 hours.